Glycosylation is a posttranslational modification of a protein, such as an antibody, which adds linear or branched chains of monosaccharide units (i.e., glycans) to the protein. Glycosylation creates a vast repertoire of structural and functional diversity on proteins. For example, the culture conditions, choice of expression systems, and the conformation of the antibodies introduce a high degree of structural and sequence heterogeneity in the glycosylation of antibodies. Glycosylation of therapeutic antibodies produced in mammalian expression systems occurs in the constant region (Fc), as well as the heavy and light chain variable regions (Fab), and can significantly modify antibody functions, such as Fc receptor binding, complement activation, and antigen binding affinity.
For antibody product development the identification of the location of the constant and variable region glycans and a complete characterization of their heterogeneity is essential. Specifically, the antibody glycosylation pattern must be controlled during biopharmaceutical production to maintain the efficacy and safety of the therapeutic. The current methods for the characterization of glycopeptides are laborious and are not capable of identifying the location of the constant and variable region glycans of an antibody, particularly when the sequence of the antibody is unknown. Therefore, more efficient methods are needed.